Construction and expression of the prM-E-△-PA3 fusion protein
Based on the baculovirus expression system, we constructed several
prM-E-△ -PA3 fragments fusing the ZIKV prM-E-△ gene
(ST-TM region deletion) and the gene for the carrier protein PA3, as
shown in Fig. 1A. In the construction strategies, signal peptides from
different sources, such as JEV and gp67 (a surface glycoprotein of
baculovirus), were considered. Three kinds of recombinant baculoviruses
(ZI-△-PA vir, ZI-JE-△-PA vir and ZI-GP-△-PA vir) were constructed and
were seeded in Sf9 cells for ZIKV-E protein detection. IFA of the ZIKV-E
protein showed positive results, with bright green fluorescence (Fig.
1B), and the cell control showed red due to Evans blue. To determine the
localization of the target protein, the supernatants and sonicated
supernatants of cell cultures were subjected to WB analysis. The results
showed that ZI-△-PA, ZI-JE-△-PA and ZI-GP-△-PA protein were secreted
into the supernatant (Fig. 1C). The IFA and WB results implied that the
prM-E-△ -PA3 (ZI-△-PA, ZI-JE-△-PA and ZI-GP-△-PA) protein
reacted with an anti-flavivirus mAb with good antigenicity. When ST-TM
was not included, the target protein was secreted.